Preclinical studies demonstrate synergism between cancer immunotherapy and local radiation, enhancing anti-tumor effects and promoting immune responses. BI1361849 (CV9202) is an active cancer ...immunotherapeutic comprising protamine-formulated, sequence-optimized mRNA encoding six non-small cell lung cancer (NSCLC)-associated antigens (NY-ESO-1, MAGE-C1, MAGE-C2, survivin, 5T4, and MUC-1), intended to induce targeted immune responses.
We describe a phase Ib clinical trial evaluating treatment with BI1361849 combined with local radiation in 26 stage IV NSCLC patients with partial response (PR)/stable disease (SD) after standard first-line therapy. Patients were stratified into three strata (1: non-squamous NSCLC, no epidermal growth factor receptor (EGFR) mutation, PR/SD after ≥4 cycles of platinum- and pemetrexed-based treatment n = 16; 2: squamous NSCLC, PR/SD after ≥4 cycles of platinum-based and non-platinum compound treatment n = 8; 3: non-squamous NSCLC, EGFR mutation, PR/SD after ≥3 and ≤ 6 months EGFR-tyrosine kinase inhibitor (TKI) treatment n = 2). Patients received intradermal BI1361849, local radiation (4 × 5 Gy), then BI1361849 until disease progression. Strata 1 and 3 also had maintenance pemetrexed or continued EGFR-TKI therapy, respectively. The primary endpoint was evaluation of safety; secondary objectives included assessment of clinical efficacy (every 6 weeks during treatment) and of immune response (on Days 1 baseline, 19 and 61).
Study treatment was well tolerated; injection site reactions and flu-like symptoms were the most common BI1361849-related adverse events. Three patients had grade 3 BI1361849-related adverse events (fatigue, pyrexia); there was one grade 3 radiation-related event (dysphagia). In comparison to baseline, immunomonitoring revealed increased BI1361849 antigen-specific immune responses in the majority of patients (84%), whereby antigen-specific antibody levels were increased in 80% and functional T cells in 40% of patients, and involvement of multiple antigen specificities was evident in 52% of patients. One patient had a partial response in combination with pemetrexed maintenance, and 46.2% achieved stable disease as best overall response. Best overall response was SD in 57.7% for target lesions.
The results support further investigation of mRNA-based immunotherapy in NSCLC including combinations with immune checkpoint inhibitors.
ClinicalTrials.gov identifier: NCT01915524 .
In mice, injection of messenger RNA (mRNA) coding for tumor-associated antigens can induce antitumor immune responses and therefore offers a broadly applicable immunotherapy approach. We injected ...intradermally protamine-stabilized mRNAs coding for Melan-A, Tyrosinase, gp100, Mage-A1, Mage-A3, and Survivin in 21 metastatic melanoma patients. In 10 patients keyhole limpet hemocyanin (KLH) was added to the vaccine. Granulocyte macrophage colony-stimulating factor was applied as an adjuvant. Endpoints were toxicity and immune responses. No adverse events more than grade II have been observed. During treatment the frequency of Foxp3+/CD4+ regulatory T cells was significantly decreased upon mRNA vaccination in peripheral blood of the patients in the KLH arm, whereas myeloid suppressor cells (CD11b+HLA-DR lo monocytes) were reduced in the patients not receiving KLH. A reproducible increase of vaccine-directed T cells was observed in 2 of 4 immunologically evaluable patients. One of 7 patients with measurable disease showed a complete response. In conclusion, we show here that direct injection of protamine-protected mRNA is feasible and safe. The significant influence of the treatment on the frequency of immunosuppressive cells, the increase of vaccine-directed T cells upon treatment in a subset of patients together with the demonstration of a complete clinical response encourage further clinical investigation of the protamine-mRNA vaccine.
Stabilized synthetic RNA oligonucleotides (ORN) and protected messenger RNA (mRNA) were recently discovered to possess an immunostimulatory capacity through their recognition by TLR 7 and 8. We ...wanted to find out whether this danger signal is capable of triggering anti‐tumor immunity when injected locally into an established tumor. Using the mouse glioma tumor cell line SMA‐560 in syngenic VM/Dk mice, we were able to show that intra‐tumor injections of protamine‐stabilized mRNA do indeed induce tumor regression and long‐term anti‐tumor immunity. Residual RNA‐injected tumors show CD8 infiltration. Distant injections of protamine‐protected mRNA and intra‐tumor injection of naked mRNA also result in anti‐tumor immunity. Although they are strong danger signals, RNA are labile molecules with a short half‐life: they do not trigger side effects such as long‐term, uncontrolled immunostimulation evidenced by splenomegaly in CpG DNA‐treated mice. In conclusion, RNA molecules are potent and safe danger signals that are relevant for active immunotherapy strategies aimed at the eradication of solid tumors.
We reported that RNA condensed on protamine is protected from RNase‐mediated degradation and can be used for vaccination. Here, we show that such complexes are also danger signals that activate mouse ...cells through a MyD88‐dependent pathway. Moreover, mRNA‐protamine complexes stimulate human blood cells. They strongly activate DC and monocytes, leading to TNF‐α and IFN‐α secretion. In addition, protamine‐RNA complexes directly activate B cells, NK cells and granulocytes. The detailed analysis of the activated cell types, the study of the cytokines released from PBMC cultured with protamine‐RNA complexes and recently published results suggest that TLR‐7 and TLR‐8 may be involved in the recognition of protamine‐stabilized RNA. Our data indicate that protamine‐stabilized RNA, which may be similar to RNA condensed in the nucleocapsids of RNA viruses, is a strong danger signal. Thus, similarly to plasmid DNA, protamine‐RNA combines antigen production and non‐specific immunostimulation. The studies presented here explain the capacity of protamine‐RNA to act as a vaccine, and pave the way towards the development of safe and efficient mRNA‐based immunotherapies.
CV9201 is an RNActive
®
-based cancer immunotherapy encoding five non-small cell lung cancer-antigens: New York esophageal squamous cell carcinoma-1, melanoma antigen family C1/C2, survivin, and ...trophoblast glycoprotein. In a phase I/IIa dose-escalation trial, 46 patients with locally advanced (
n
= 7) or metastatic (
n
= 39) NSCLC and at least stable disease after first-line treatment received five intradermal CV9201 injections (400–1600 µg of mRNA). The primary objective of the trial was to assess safety. Secondary objectives included assessment of antibody and ex vivo T cell responses against the five antigens, and changes in immune cell populations. All CV9201 dose levels were well-tolerated and the recommended dose for phase IIa was 1600 µg. Most AEs were mild-to-moderate injection site reactions and flu-like symptoms. Three (7%) patients had grade 3 related AEs. No related grade 4/5 or related serious AEs occurred. In phase IIa, antigen-specific immune responses against ≥ 1 antigen were detected in 63% of evaluable patients after treatment. The frequency of activated IgD
+
CD38
hi
B cells increased > twofold in 18/30 (60%) evaluable patients. 9/29 (31%) evaluable patients in phase IIa had stable disease and 20/29 (69%) had progressive disease. Median progression-free and overall survival were 5.0 months (95% CI 1.8–6.3) and 10.8 months (8.1–16.7) from first administration, respectively. Two- and 3-year survival rates were 26.7% and 20.7%, respectively. CV9201 was well-tolerated and immune responses could be detected after treatment supporting further clinical investigation.
Protein‐ and peptide‐based tumor vaccines depend on strong adjuvants to induce potent immune responses. Here, we demonstrated that a recently developed novel adjuvant based on a non‐coding, ...long‐chain RNA molecule, termed RNAdjuvant®, profoundly increased immunogenicity of both antigen formats. RNAdjuvant® induced balanced, long‐lasting immune responses that resulted in a strong anti‐tumor activity. A direct comparison to Poly(I:C) showed superior efficacy of our adjuvant to enhance antigen‐specific multifunctional CD8+ T‐cell responses and mediate anti‐tumor responses induced by peptide derived from HPV‐16 E7 protein in the syngeneic TC‐1 tumor, a murine model of human HPV‐induced cervical cancer. Moreover, the adjuvant was able to induce functional memory responses that mediated complete tumor remission. Despite its remarkable immunostimulatory activity, our RNA‐based adjuvant exhibited an excellent pre‐clinical safety profile. It acted only locally at the injection site where it elicited a transient but strong up‐regulation of pro‐inflammatory and anti‐viral cytokines as well as cytoplasmic RNA sensors without systemic cytokine release. This was followed by the activation of immune cells in the draining lymph nodes. Our data indicate that our RNA‐based adjuvant is a safe and potent immunostimulator that may profoundly improve the efficacy of a variety of cancer vaccines.
What's new?
While adjuvants can enhance immune responses induced by peptide and protein vaccines, in cancer immunotherapy, they fail to trigger the balanced, long‐lasting immunity that is needed for anti‐tumor protection. That limitation may be overcome with the development of synthetic RNA molecules as adjuvants, according to the present study. In a TC‐1 tumor mouse model, RNAdjuvant®, a novel synthetic RNA with a polymeric carrier, significantly enhanced immune responses associated with the administration of peptide vaccine. Profound anti‐tumor effects were observed. The adjuvant acted locally, at the site of injection, without inducing systemic cytokine release.
A novel, disruptive vaccination technology Kallen, Karl-Josef; Heidenreich, Regina; Schnee, Margit ...
Human vaccines & immunotherapeutics,
10/2013, Letnik:
9, Številka:
10
Journal Article
Recenzirano
Odprti dostop
Nucleotide based vaccines represent an enticing, novel approach to vaccination. We have developed a novel immunization technology, RNActive
®
vaccines, that have two important characteristics: mRNA ...molecules are used whose protein expression capacity has been enhanced by 4 to 5 orders of magnitude by modifications of the nucleotide sequence with the naturally occurring nucleotides A (adenosine), G (guanosine), C (cytosine), U (uridine) that do not affect the primary amino acid sequence. Second, they are complexed with protamine and thus activate the immune system by involvement of toll-like receptor (TLR) 7. Essentially, this bestows self-adjuvant activity on RNActive
®
vaccines. RNActive
®
vaccines induce strong, balanced immune responses comprising humoral and cellular responses, effector and memory responses as well as activation of important subpopulations of immune cells, such as Th1 and Th2 cells. Pre-germinal center and germinal center B cells were detected in human patients upon vaccination. RNActive
®
vaccines successfully protect against lethal challenges with a variety of different influenza strains in preclinical models. Anti-tumor activity was observed preclinically under therapeutic as well as prophylactic conditions. Initial clinical experiences suggest that the preclinical immunogenicity of RNActive
®
could be successfully translated to humans.
CV9103 is a prostate-cancer vaccine containing self-adjuvanted mRNA (RNActive®) encoding the antigens PSA, PSCA, PSMA, and STEAP1. This phase I/IIa study evaluated safety and immunogenicity of CV9103 ...in patients with advanced castration-resistant prostate-cancer.
44 Patients received up to 5 intra-dermal vaccinations. Three dose levels of total mRNA were tested in Phase I in cohorts of 3-6 patients to determine a recommended dose. In phase II, 32 additional patients were treated at the recommended dose. The primary endpoint was safety and tolerability, the secondary endpoint was induction of antigen specific immune responses monitored at baseline and at weeks 5, 9 and 17.
The most frequent adverse events were grade 1/2 injection site erythema, injection site reactions, fatigue, pyrexia, chills and influenza-like illness. Possibly treatment related urinary retention occurred in 3 patients. The recommended dose was 1280 μg. A total of 26/33 evaluable patients treated at 1280 μg developed an immune response, directed against multiple antigens in 15 out of 33 patients. One patient showed a confirmed PSA response. In the subgroup of 36 metastatic patients, the Kaplan-Meier estimate of median overall survival was 31.4 months 95 % CI: 21.2; n.a.
The self-adjuvanted RNActive® vaccine CV9103 was well tolerated and immunogenic. The technology is a versatile, fast and cost-effective platform allowing for creation of vaccines. The follow-up vaccine CV9104 including the additional antigens prostatic acid phosphatase (PAP) and Muc1 is currently being tested in a randomized phase IIb trial to assess the clinical benefit induced by this new vaccination approach.
EU Clinical Trials Register: EudraCT number 2008-003967-37, registered 27 Jan 2009.
Vaccination against tumor antigens has been shown to be a safe and efficacious prophylactic and therapeutic antitumor treatment in many animal models. Clinical studies in humans indicate that ...specific immunotherapy can also result in clinical benefits. The active pharmaceutical ingredient in such vaccines can be DNA, RNA, protein, or peptide and can be administered naked, encapsulated, or after delivery in vitro into cells that are then adoptively transferred. One of the easiest, most versatile and theoretically safest technologies relies on the direct injection of naked messenger RNA (mRNA) that code for tumor antigens. We and others have shown in mice that intradermal application of naked mRNA results in protein expression and the development of an immune response. We used this protocol to vaccinate 15 melanoma patients. For each patient a growing metastasis was removed, total RNA was extracted, reverse-transcribed, amplified, and cloned. Libraries of cDNA were transcribed to produce unlimited amounts of copy mRNA. Autologous preparations were applied intradermally in combination with granulocyte macrophage colony-stimulating factor as adjuvant. We demonstrate here that such treatment is feasible and safe (phase 1 criteria). Furthermore, an increase in antitumor humoral immune response was seen in some patients. However, a demonstration of clinical effectiveness of direct injection of copy mRNA for antitumor immunotherapy was not shown in this study and must be evaluated in subsequent trials.
Abstract
CV8102 comprises a single-stranded non-coding RNA complexed with a cationic peptide. It acts as an agonist to TLR-7/-8 and RIG-I (Ziegler 2018) to stimulate the innate and adaptive immune ...system. CV8102 was shown to induce an upregulation of inflammatory cytokines, chemokines and IFN-γ related genes at the injection site along with an activation of T, NK, NKT and migratory dendritic cells in the draining lymph nodes (Heidenreich 2015). Intratumoral (IT) CV8102 demonstrated dose-dependent anti-tumor activity and synergized with systemic PD-1 inhibition in preclinical models.
Methods This Phase I study investigates IT CV8102 as single agent and in combination with systemic anti-PD-1 antibodies (as per product label). Patients (pts) with advanced inoperable melanoma (MEL), cutaneous/head and neck squamous cell or adenoid cystic carcinoma (cSCC, SCCHN, ACC) are eligible for single agent CV8102, pts with MEL and SCCHN who did not respond or slowly progressed on anti-PD-1 therapy are eligible for the combination. CV8102 is administered for up to 8 IT injections into a single accessible tumor lesion over a 12-week period. A Bayesian logistic regression model with overdose control is used for the dose escalation parts. Response is assessed by RECIST 1.1/irRECIST (injected and non-injected target lesions). Pre- and on-treatment samples are collected for biomarker analyses.
Results A total of 20 pts have been treated with either CV8102 alone (N=15: 6 MEL, 2 SCCHN, 5 ACC, 2 cSCC) or CV8102 in combination with anti-PD-1 (N=5: 4 MEL, 1 SCCHN). Dose cohorts with doses up to 150 µg (CV8102 alone) and 100µg (CV8102+anti-PD-1) have been completed. Most common AEs were mild to moderate flu-like symptoms and injection site reactions. No dose limiting toxicities (DLT) were observed during the DLT period of the first two weeks of treatment. 12 pts treated with CV8102 alone were evaluable for response assessment. 1 MEL pt treated at 150µg experienced a complete regression of injected and non-injected lesions. This patient also experienced a marked increase of IL-6 and CRP at 6 and 24 hours after the first injection, respectively. 7 pts achieved stable disease, with two pts treated at 100 µg (SCCHN pt) and 200µg (MEL pt who had developed acquired resistance to previous anti-PD-1 therapy) showing regression of non-injected lymph node lesions. Dose escalation parts are continuing, updated safety and efficacy results will be presented.
Conclusion Intratumoral single agent CV8102 appears well tolerated and showed preliminary evidence of clinical efficacy with shrinkage of injected and non-injected lesions.
Citation Format: Thomas Eigentler, Juergen Krauss, Jutta Schreiber, Carsten Weishaupt, Patrick Terheyden, Lucie Heinzerling, Peter Mohr, Benjamin Weide, Ralf Gutzmer, Juergen C. Becker, Felix Kiecker, Angelika Daehling, Fatma Funkner, Regina Heidenreich, Sarah-Katharina Kays, Ute Klinkhardt, Birgit Scheel, Oliver Schoenborn-Kellenberger, Tobias Seibel, Claudia Stosnach, Tanja Strack, Ulrike Gnad-Vogt. Intratumoral RNA-based TLR-7/-8 and RIG-I agonist CV8102 alone and in combination with anti-PD-1 in a Phase I dose-escalation and expansion trial in patients with advanced solid tumors abstract. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr LB-021.
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